The routine use of hematopoietic stem cell (HSC) transplantation as an effective treatment of hematologic and malignant diseases is hampered by the limited availability of HSC-enriched cell populations In addition, the success rate of HSC transplant therapies could be significantly increased if large quantities of transplant material with a high HSC content could be reliably generated. An increased HSC supply would also greatly stimulate research activities based on gene therapy of the hematopoietic system. The long term goal of this project is to address these limitations by developing a well defined culture system capable of specifically expanding an HSC-enriched progenitor population while simultaneously maintaining or increasing the HSC percentage. In most hematopoietic culture and expansion schemes, expansion and lineage differentiation proceed in parallel and HSC expansion per se is limited or non-existent. Recent studies suggest that in addition to manipulation of the cytokine milieu, it may be possible to direct the differentiation vs. proliferation activities of hematopoietic populations by controlling the nature and levels of glycosaminoglycan (GAG) substances to which the cells are exposed. Our previous work showed that this approach can be used to generate very high yield expansions of umbilical cord blood progenitors while simultaneously maintaining and enriching the culture with CD34+ cells. In the proposed studies, we will: (1) fully characterize and refine our culture system; (2) characterize the expanded cell populations with regard to HSC attributes; (3) develop and validate mechanistic mathematical models of GAG mediated hematopoietic control; and (4) use these results to design and optimize a perfusion bioreactor system based upon the polysaccharide surfaces. The bioreactor operating parameters will be optimized with the goal of producing, high yield expansion of cord blood progenitor populations. These studies will provide both fundamental knowledge about the poorly understood roles of various matrix polysaccharides in hematopoiesis, as well as the technology base for ultimately attaining specific non-differentiative expansion of pluripotent HSCs.